( activity and production of ...)High-Efficiency Site-Directed Mutagenesis of Large Plasmid Vectors

activity and production of infectious virus (data not shown).

Table 1
Successful Mutagenesis of a Large Plasmid with the QuikChange XL Kit

DNA Template
(kb)

Extension Time
(minutes/kb)

Number of Transformants^a

QuikChange kit

QuikChange XL kit

10.1

1

0

32

^aReactions were carried out as described in the text and kit manuals. Values represent mean number of transformants from 3 plates.

Small-to-moderately sized plasmid DNA templates can be efficiently mutagenized with either QuikChange kit⁴. For such targets, the QuikChange XL kit permits the use of reduced reaction times. For example, the 4.5-kb kit control was efficiently replicated with the QuikChange XL kit using 0.5 min/kb extension times rather the 2 minute/kb extension times recommended for the standard QuikChange kit (data not shown). In these experiments, the average mutation frequency for the kit control was 98%, indicating that QuikChange XL kit retains the high mutagenesis efficiency of the QuikChange site-directed mutagenesis method.

Conclusions

We have introduced the QuikChange XL site-directed mutagenesis kit to improve the efficiency of mutagenizing long or difficult-to-mutagenize plasmid DNA templates using this single day mutagenesis method. The QuikChange XL kit features components specifically designed for more efficient DNA replication and bacterial t
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